Protein-protein interactions among transcription factors play critical roles in the regulation of gene expression in normal and disease states. This application proposes to study protein-protein interactions mediated by Cys2His2 zinc finger proteins, the most abundant class of transcription factors in humans. In particular, we are focusing our studies on dimerization zinc finger (DZF) domains found in members of the Ikaros family of transcription factors and related homologues. DZF domains provide a model system for studying Cys2His2 zinc finger-mediated protein-protein interactions. The long-term goals of this proposal are: to gain a detailed biophysical understanding of how Cys2His2 zinc fingers mediate specific interactions with a wide variety of different proteins; to utilize this knowledge to devise strategies to enhance, disrupt, or predict such interactions; and to engineer synthetic zinc fingers with novel interaction specificities that could serve as useful modules for creating artificial regulatory circuits with potential applications in biotechnology and therapeutics. The specific aims of this application are: 1) to define and model the Ikaros, Pegasus, and Hunchback DZF domain interfaces using mutational analysis, genetic selection methods, and molecular graphics tools; 2) to determine high-resolution structures of dimeric Ikaros, Pegasus, and Hunchback DZF domain protein-protein complexes using X-ray crystallography; 3) to use genetic selection to identify synthetic, obligate heterodimeric DZF domains and to test the use of DZF domains as "interaction modules" for assembling heterologous protein complexes in eukaryotic cells.